Adhesion G protein-coupled receptors are activated by exposure of a cryptic tethered agonist

The large class of adhesion G protein-coupled receptors (aGPCRs) bind extracellular matrix or neighboring cell-surface ligands to regulate organ and tissue development through an unknown activation mechanism. We examined aGPCR activation using two prototypical aGPCRs, GPR56 and GPR110. Active dissociation of the noncovalently bound GPR56 or GPR110 extracellular domains (ECDs) from the respective seven-transmembrane (7TM) domains relieved an inhibitory influence and permitted both receptors to activate defined G protein subtypes. After ECD displacement, the newly revealed short N-terminal stalk regions of the 7TM domains were found to be essential for G protein activation. Synthetic peptides comprising these stalks potently activated GPR56 or GPR110 in vitro or in cells, demonstrating that the stalks comprise a tethered agonist that was encrypted within the ECD. Establishment of an aGPCR activation mechanism provides a rational platform for the development of aGPCR synthetic modulators that could find clinical utility toward aGPCR-directed disease. Significance Adhesion G protein-coupled receptors (GPCRs) regulate tissue development and cancer progression. Intact adhesion GPCRs are unique two-protomer receptors that undergo biosynthetic self-proteolyzation at a conserved site between the extracellular domain and the seven transmembrane (7TM) domain. We demonstrate the activation mechanism of adhesion GPCRs. The 7TM domain N-terminal stalk lies encrypted within the noncovalently bound extracellular domain. When the extracellular domain is dissociated, perhaps naturally by ligand action, the 7TM stalk is revealed. We demonstrate that the stalk acts as a tethered agonist of the 7TM to facilitate direct activation of heterotrimeric G proteins. Synthetic peptides comprising adhesion GPCR 7TM domain stalks are sufficient receptor agonists, raising the possibility for the development of adhesion GPCR synthetic peptide modulators.